Sex-dependent differences in cardiovascular disease (CVD) susceptibility and progression may be estrogen-mediated, but the genetic pathways regulating estrogen cardioprotection have yet to be elucidated. Receptor Activity Modifying Proteins, or RAMPs, are transmembrane accessory proteins that are known to interact with G-protein coupled receptors (GPCRs) and play a role in receptor trafficking and ligand specificity. RAMP3 is unique in that it is transcriptionally induced by estrogen and CVD. Also, unlike other RAMP family members, RAMP3 is known for its ability to alter GPCR recycling to the plasma membrane through a PDZ motif-dependent association with NSF. GPER1 is a novel cardioprotective estrogen-binding GPCR that is present in increased levels in females and in CVD. Our preliminary findings have shown that genetic loss of RAMP3 leads to reduced levels of GPER1 at the plasma membrane of cardiac cells in a sex-dependent fashion, providing evidence for an interaction of RAMP3 with GPER1. We will investigate the potential interaction between RAMP3 and GPER1 in vitro using transfected cells to determine whether RAMP3 can alter GPER1 recycling to the plasma membrane. In addition we will perform in vivo studies to determine whether pharmacological activation of GPER1 signaling affects the cardiovascular phenotype of RAMP3-/- male and female mice on a genetic background of heart failure. Results from this proposal will elucidate the role of RAMP3 and GPER1 in sex-dependent cardioprotection and have the potential of identifying novel therapeutic targets for gender-tailored treatment of CVD.